Coding and decoding method for determined nucleic acid sequence

A nucleic acid sequence and coding method technology, applied in biochemical equipment and methods, microbial determination/inspection, etc., can solve the problems of large number of samples and low sample throughput, and achieves simple method, simplified operation process, and wide application. Effect

Inactive Publication Date: 2008-06-18
SOUTHEAST UNIV
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Problems solved by technology

[0005] The invention provides a method for encoding and decoding nucleic acid sequences to be tested that can effectively solve the contradiction between the existing high-energy DNA sequence determination technology and the low throughput of samples measured by users but the large number of samples. The invention helps to reduce the cost of determination , which has the advantage of being simple

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  • Coding and decoding method for determined nucleic acid sequence

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example 1

[0026] Example 1: High-throughput analysis of DNA sequences encoding different samples.

[0027] Cut different sample DNA sequences with enzymes (or sonicate) into fragments with a size of 50-100 bases, and connect these fragmented nucleic acid sequences with a pair of linkers under the action of ligase (one of which is a universal linker) One sub-oligonucleotide sequence is completely complementary to the sequence of the amplification primer, and the other is a unique known sequence code corresponding to different samples and a fragment of the same oligonucleotide sequence as the sequencing primer.

[0028] The fragmented nucleic acid sequence connected by these tethers and the complementary sequence of the fixed linker are transferred to microbeads for emulsion parallel PCR reaction to amplify the fragmented whole human genome. These microbeads are immobilized on the plate substrate, and sequencing templates of DNA sequences of different samples are obtained by enzyme digest...

example 2

[0030] Example 2: Expression analysis of mRNA sequences encoding different sources.

[0031] Step 1: Extract cellular mRNA from various sources, and use magnetic beads to immobilize 30 T nucleotide chains to purify mRNA by hybridizing mRNA.

[0032] Step 2: respectively reverse transcribe into cDNA, and use RNaseH, DNA polymerase, etc. to synthesize the second strand.

[0033] Step 3: Digest the above double-stranded nucleic acid sequence with an endonuclease (Sau 3AI) that recognizes four bases, and wash the magnetic beads with a buffer.

[0034] Step 4: under the action of ligase, add the above-mentioned digested double-stranded nucleic acid sequence to the linker primer 3 (the primer has a recognition site of endonuclease Acu I).

[0035] Step 5: Digest the product of step 4 with Acu I. A cDNA library of adapter primer 3 followed by 16 unknown sequences to be tested was obtained.

[0036] Step 6: The product of step 5 is connected with a linker primer 4 to complete the e...

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Abstract

The invention discloses a coding method of a nucleic acid sequence to be detected which is a method of simultaneously analyzing high throughputs of nucleic acid sequences of different samples or different nucleic sequences of one sample and in particular relates to a coding and decoding method of the nucleic acid sequence to be detected. The scheme of the invention is to respectively cut every nucleic sequence to be detected into nucleic acid fragments and then respectively connect one connexon with the two ends of the nucleic acid fragment to form templates. One connexon consists of an amplified (complementary to a sequencing primer) sequence fragment and a section of sequence codes. And the sequence codes on the nucleic fragment of the same nucleic sequence are same. Thus, the nucleic acid sequences of the codes are formed. The sequence codes consists of four bases of A, T, C or G and are obtained through a synthetic method. The invention can solve the contradict between the present high-energy DNA sequence detection technology and the low throughput of samples to be detected by users with multiple samples and lower the detection cost. The method is simple.

Description

technical field [0001] The invention is a method for realizing high-throughput simultaneous analysis of nucleic acid sequences of different samples or different nucleic acid sequences of the same sample, and in particular relates to a method for encoding and decoding nucleic acid sequences to be tested. technical background [0002] With the development and completion of the Human Genome Project and various model organism genome projects, human beings have entered the post-gene era, which has had a huge impact on contemporary biological research and medical research, and the related disciplines of molecular biology have been rapidly developed. develop. It will become possible to understand the differences of life, the law of disease occurrence and development, and the interaction between drugs and living organisms at the genetic level. As far as gene sequence analysis is concerned, the focus of the post-genome era has shifted from whole-genome sequence determination to the ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 肖鹏峰陆祖宏李燕强潘志强唐静
Owner SOUTHEAST UNIV
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